The following description includes information that may be useful in understanding the present invention. It is not an admission that any of the information provided herein is prior art to the presently claimed invention, nor that any of the publications specifically or implicitly referenced are prior art to that invention.
The exendins are peptides that are found in the salivary secretions of the Gila monster and the Mexican Bearded Lizard, reptiles that are endogenous to Arizona and Northern Mexico. Exendin-3 (SEQ ID NO: 1) is present in the salivary secretions of Heloderma horridum (Mexican Beaded Lizard), and exendin-4 (SEQ ID NO: 2) is present in the salivary secretions of Heloderm suspectum (Gila monster)(Eng et al, J. Biol. Chem., 265:20259-62 (1990); Eng et al, J. Biol. Chem., 267:7402-05 (1992). The amino acid sequence of exendin-3 is shown in FIG. 1. The amino acid sequence of exendin-4 is shown in FIG. 2. Exendin-4 was first thought to be a (potentially toxic) component of the venom. It now appears that exendin-4 is devoid of toxicity, and that it instead is made in salivary glands in the Gila monster.
The exendins have some sequence similarity to several members of the glucagon-like peptide family, with the highest homology, 53%, being to GLP-1[7-36]NH2 (SEQ ID NO: 3) (Goke et al, J. Biol. Chem., 268:19650-55 (1993). GLP-1[7-36]NH2, also sometimes referred to as proglucagon[78-107] or simply “GLP-1”, has an insulinotropic effect, stimulating insulin secretion from pancreatic beta-cells; GLP-1 has also been reported to inhibit glucagon secretion from pancreatic alpha-cells (Ørsov et al, Diabetes, 42:658-61 (1993); D'Alessio et al, J. Clin. Invest., 97:133-38 (1996)). GLP-1 has been reported to inhibit gastric emptying (Willms et al, J. Clin. Endocrinol. Metab., 81(1):327-32 (1996); Wettergren et al, Dig. Dis. Sci., 38(4):665-73 (1993)), and gastric acid secretion (Schjoldager et al, Dig. Dis. Sci., 34(5):703-8 (1989); O'Halloran et al, J. Endocrinol., 126(1):169-73 (1990); Wettergren et al, Dig. Dis. Sci., 38(4):665-73 (1993)). GLP-1[7-37], which has an additional glycine residue at its carboxy terminus, is reported to stimulate insulin secretion in humans (Ørsov et al, Diabetes, 42:658-61 (1993)). Other reports relate to the inhibition of glucagon secretion (Creutzfeldt et al, “Glucagonostatic actions and reduction of fasting hyperglycemia by exogenous glucagon-like peptide I(7-36) amide in Type 1 diabetic patients,” Diabetes Care, 19(6):580-6 (1996)), and a purported role in appetite control (Turton et al, “A role for glucagon-like peptide-1 in the central regulation of feeding,” Nature, 379(6560):69-72 (1996)). A transmembrane G-protein adenylate-cyclase-coupled receptor, said to be responsible at least in part for the insulinotropic effect of GLP-1, has reportedly been cloned from a beta-cell line (Thorens, Proc. Natl. Acad. Sci. USA, 89:8641-45 (1992)). GLP-1 has been the focus of significant investigation in recent years due to its reported action on the amplification of stimulated insulin production (Byrne M M, Goke B. Lessons from human studies with glucagon-like peptide-1: Potential of the gut hormone for clinical use. In: Fehmann H C, Goke B. Insulinotropic Gut Hormone Glucagon-Like Peptide 1. Basel, Switzerland: Karger, 1997:219-33).
GLP-1 has also been reported to restore islet glucose sensitivity in aging rats, restoring their glucose tolerance to that of younger rats (Egan J M, et al., Diabetologia 1997 June; 40(Suppl 1):A130). However, the short duration of biological action of GLP-1 in vivo is one feature of the peptide that has hampered its development as a therapeutic agent. Various methods have been tried to prolong the half-life of GLP-1 or GLP-1 (7-37), including attempts to alter their amino acid sequences and to deliver them using certain formulations (see, e.g., European Patent Application, entitled “Prolonged Delivery of Peptides,” by Darley, et al., publication number 0 619 322 A2, regarding the inclusion of polyethylene glycol in formulations containing GLP-1 (7-37)).
Pharmacological studies have led to reports that exendin-4 can act at GLP-1 receptors in vitro on certain insulin-secreting cells, at dispersed acinar cells from guinea pig pancreas, and at parietal cells from stomach; the peptide is also reported to stimulate somatostatin release and inhibit gastrin release in isolated stomachs (Goke, et al., J. Biol. Chem. 268:19650-55, 1993; Schepp, et al., Eur. J. Pharmacol., 69:183-91, 1994; Eissele, et al., Life Sci., 55:629-34, 1994). Exendin-3 and exendin-4 were reportedly found to stimulate cAMP production in, and amylase release from, pancreatic acinar cells (Malhotra, R., et al., Regulatory Peptides, 41:149-56, 1992; Raulman, et al., J. Biol. Chem. 267:21432-37, 1992; Singh, et al., Regul Pept. 53:47-59, 1994). Exendin-4 has a significantly longer duration of action than GLP-1. For example, in one experiment, glucose lowering by exendin-4 in diabetic mice was reported to persist for several hours, and, depending on dose, for up to 24 hours (Eng, J. Prolonged effect of exendin-4 on hyperglycemia of db/db mice, Diabetes 1996 May; 45(Suppl 2):152A (abstract 554)). Based on their insulinotropic activities, the use of exendin-3 and exendin-4 for the treatment of diabetes mellitus and the prevention of hyperglycemia has been proposed (Eng, U.S. Pat. No. 5,424,286).
The results of an investigation which showed that exendins are not the species homolog of mammalian GLP-1 was reported by Chen and Drucker who cloned the exendin gene from the Gila monster (J. Biol. Chem. 272(7):4108-15 (1997)). The observation that the Gila monster also has separate genes for proglucagons (from which GLP-1 is processed), that are more similar to mammalian proglucagon than exendin, indicated that exendins are not merely species homologs of GLP-1.
Methods for regulating gastrointestinal motility using exendin agonists are described in U.S. Pat. No. 6,858,576 (i.e., based on U.S. application Ser. No. 08/908,867 filed Aug. 8, 1997, which is a continuation-in-part of U.S. application Ser. No. 08/694,954 filed Aug. 8, 1996). Methods for reducing food intake using exendin agonists are described in U.S. Pat. No. 6,956,026 (i.e., based on U.S. application Ser. No. 09/003,869, filed Jan. 7, 1998, which claims the benefit of U.S. Application Nos. 60/034,905 filed Jan. 7, 1997, 60/055,404 filed Aug. 7, 1997, 60/065,442 filed Nov. 14, 1997, and 60/066,029 filed Nov. 14, 1997.
Novel exendin agonist compounds are described in WO 99/07404 (i.e., PCT/US98/16387 filed Aug. 6, 1998, which claims the benefit of U.S. Patent Application Ser. No. 60/055,404, filed Aug. 8, 1997). Other novel exendin agonists are described in WO 99/25727 (i.e., PCT/US98/24210, filed Nov. 13, 1998, which claims the benefit of U.S. Provisional Application No. 60/065,442 filed Nov. 14, 1997). Still other novel exendin agonists are described in WO 99/25728 (i.e., PCT/US98/24273, filed Nov. 13, 1998, which claims the benefit of U.S. Provisional Application No. 60/066,029 filed Nov. 14, 1997). Other recent advances in exendin related technology are described in WO 99/40788 (i.e., which claims priority to U.S. Application No. 60/075,122, filed Feb. 13, 1998); and in WO 00/41546, and WO 00/41548 (i.e., which claim priority to U.S. Application No. 60/116,380, filed Jan. 14, 1998).
Polyethylene glycol (PEG) modification of therapeutic peptides and proteins may yield both advantages and disadvantages. While PEG modification may lead to improved circulation time, reduced antigenicity and immunogenicity, improved solubility, resistance to proteolysis, improved bioavailability, reduced toxicity, improved stability, and easier formulation of peptides (See, Francis et al., International Journal of Hematology, 68:1-18, 1998) problems with PEGylation in most cases is substantial reduction in bioactivity. Id. In addition, most methods involve use of linkers that have several types of adverse effects including immunogenicity, instability, toxicity, and reactivity. Id.
Modified exendins and exendin agonists and related formulations, dosage formulations, and methods that solve these problems and that are useful in the delivery of therapeutically effective amounts of exendins and exendin agonists are described and claimed herein.
The contents of the above-identified articles, patents, and patent applications, and all other documents mentioned or cited herein, are hereby incorporated by reference in their entirety. The inventors reserve the right to physically incorporate into this application any and all materials and information from any such articles, patents, patent applications, or other documents mentioned or cited herein.